Heteroduplex mobility and sequence analyses for assessment of variability of Zucchini yellow mosaic virus

Citation
Ss. Lin et al., Heteroduplex mobility and sequence analyses for assessment of variability of Zucchini yellow mosaic virus, PHYTOPATHOL, 90(3), 2000, pp. 228-235
Citations number
45
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Plant Sciences
Journal title
PHYTOPATHOLOGY
ISSN journal
0031-949X → ACNP
Volume
90
Issue
3
Year of publication
2000
Pages
228 - 235
Database
ISI
SICI code
0031-949X(200003)90:3<228:HMASAF>2.0.ZU;2-C
Abstract
A heteroduplex mobility assay (HMA) was used to analyze the variability amo ng five isolates of Zucchini yellow mosaic virus (ZYMV; TW-TC1, TW-CY2, TW- TN3, TW-TNML1, and TW-NT1) collected from cucurbit fields in different area s of Taiwan. A cDNA fragment of 760 bp covering the variable region of the N terminal half of the coat protein (CP) gene was amplified by reverse tran scription-polymerase chain reaction (RT-PCR) and subsequently subjected to HMA analysis for sequence variation. When TW-NTI combined with any of the o ther Taiwan isolates, the heteroduplexes obtained migrated much more slowly than did the heteroduplexes obtained in combinations among the other four Taiwan isolates, indicating that TW-TC1, TW-CY2, TW-TN3, and TW-TNML1 share a high degree of sequence homology, while the TW-NT1 isolate is more disti nct. The complete nucleotide sequences of the CP genes and the 3' noncoding regions of the five isolates were determined from PT-PCR-derived cDNA clon es. A phylogenetic tree derived from the actual sequences of the 760-bp fra gments of the five Taiwan and another six ZYMV isolates from different geog raphic areas revealed four genotypes. TW-TNML1, TW-TC1, TWC-Y2, and TW-TN3 were in genotype I, while TW-NT1 and U.S, isolates were in genotype II. The Singapore and Reunion Island isolates were separated into genotypes III an d IV, respectively. Comparison of the CP genes of the five Taiwan isolates indicated that they share 92.8 to 98.7% nucleotide identities and 96.4 to 9 9.3% amino acid identities. The amino acid positions 73, 102, 109, and 149 of the CP gene, where lysine, serine, arginine, and aspartic acid reside, r espectively, were uniquely conserved for genotype I Taiwan isolates. Thus, results of HMA agreed well with those of phylogenetic analysis based on the sequence data of the five Taiwan ZYMV isolates. These five ZYMV isolates o f known sequence can be used as reference strains for HMA to analyze the va riability of ZYMV in Taiwan.